Splicing factors Sf3A2 and Prp31 have direct roles in mitotic chromosome segregation

Several studies have shown that RNAi-mediated depletion of splicing factors (SFs) results in mitotic abnormalities. However, it is currently unclear whether these abnormalities reflect defective splicing of specific pre-mRNAs or a direct role of the SFs in mitosis. Here, we show that two highly conserved SFs, Sf3A2 and Prp31, are required for chromosome segregation in both Drosophila and human cells. Injections of anti-Sf3A2 and anti-Prp31 antibodies into Drosophila embryos disrupt mitotic division within 1 min, arguing strongly against a splicing-related mitotic function of these factors. We demonstrate that both SFs bind spindle microtubules (MTs) and the Ndc80 complex, which in Sf3A2- and Prp31-depleted cells is not tightly associated with the kinetochores; in HeLa cells the Ndc80/HEC1-SF interaction is restricted to the M phase. These results indicate that Sf3A2 and Prp31 directly regulate interactions among kinetochores, spindle microtubules and the Ndc80 complex in both Drosophila and human cells.

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Meiotic CENP-C is a shepherd: bridging the space between the centromere and the kinetochore in time and space

Essays in Biochemistry ◽

10.1042/ebc20190080 ◽

2020 ◽

Vol 64 (2) ◽

pp. 251-261

Author(s):

Jessica E. Fellmeth ◽

Kim S. McKim

Keyword(s):

Chromosome Segregation ◽

New Technologies ◽

Early Prophase ◽

Unique Role ◽

Kinetochore Assembly ◽

M Phase ◽

In Vivo Analysis ◽

Meiosis I

Abstract While many of the proteins involved in the mitotic centromere and kinetochore are conserved in meiosis, they often gain a novel function due to the unique needs of homolog segregation during meiosis I (MI). CENP-C is a critical component of the centromere for kinetochore assembly in mitosis. Recent work, however, has highlighted the unique features of meiotic CENP-C. Centromere establishment and stability require CENP-C loading at the centromere for CENP-A function. Pre-meiotic loading of proteins necessary for homolog recombination as well as cohesion also rely on CENP-C, as do the main scaffolding components of the kinetochore. Much of this work relies on new technologies that enable in vivo analysis of meiosis like never before. Here, we strive to highlight the unique role of this highly conserved centromere protein that loads on to centromeres prior to M-phase onset, but continues to perform critical functions through chromosome segregation. CENP-C is not merely a structural link between the centromere and the kinetochore, but also a functional one joining the processes of early prophase homolog synapsis to late metaphase kinetochore assembly and signaling.

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Overexpression of SKA Complex Is Associated With Poor Prognosis in Gliomas

Frontiers in Neurology ◽

10.3389/fneur.2021.755681 ◽

2022 ◽

Vol 12 ◽

Author(s):

Shoukai Yu

Keyword(s):

Chromosome Segregation ◽

Poor Prognosis ◽

Nuclear Division ◽

Prognostic Significance ◽

Mrna Levels ◽

Ppi Network ◽

Hub Genes ◽

Protein Protein Interaction ◽

Spindle Microtubules

The spindle and kinetochore-associated complex is composed of three members: SKA1, SKA2, and SKA3. It is necessary for stabilizing spindle microtubules attaching to kinetochore (KT) in the middle stage of mitosis. The SKA complex is associated with poor prognosis in several human cancers. However, the role of SKA complex in rare malignant diseases, such as gliomas, has not been fully investigated. We investigated several databases, including Oncomine, UALCAN, and cBioPortal to explore the expression profile and prognostic significance of SKA complex in patients with gliomas. Gene ontology and Kyoto Encyclopedia of Genes and Genome pathways were used to analyze the potential enriched pathways. The genes co-expressed with SKA complex were identified and used for developing a protein-protein interaction (PPI) network using the STRING database. We found a significant overexpression of the mRNA levels of SKA1, SKA2, and SKA3 in patients with glioma patients. Higher expression of SKA1 and SKA3, but not SKA2, was significantly correlated with shorter overall survival of patients with glioma. In glioma, SKA complex was found to be involved in nuclear division, chromosome segregation, and DNA replication. The results of PPI network identified 10 hub genes (CCNB2, UBE2C, BUB1B, TPX2, CCNA2, CCNB1, MELK, TOP2A, PBK, and KIF11), all of which were overexpressed and negatively associated with prognosis of patients with glioma. In conclusion, our study sheds new insights into the biological role and prognostic significance of SKA complex in glioma.

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Phylogenetic and Expression Analysis of CENH3 and APOLLO Genes in Sexual and Apomictic Boechera Species

10.20944/preprints202101.0320.v1 ◽

2021 ◽

Author(s):

Evgeny Bakin ◽

Fatih Sezer ◽

Irem Kilic ◽

Aslıhan Özbilen ◽

Mike Rayko ◽

...

Keyword(s):

Chromosome Segregation ◽

Genetic Regulation ◽

Single Copy ◽

Egg Cell ◽

Expression Levels ◽

Histone H3 Variant ◽

Before And After ◽

Spindle Microtubules ◽

Mitosis And Meiosis

Apomictic plants (reproducing via asexual seeds), unlike sexual individuals, avoid meiosis and egg cell fertilization. Consequently, apomixis is very important for fixing maternal genotypes in the next plant generations. Despite the progress in the study of apomixis, molecular and genetic regulation of the latter remains poorly understood. So far APOLLO (Aspartate Glutamate Aspartate Aspartate histidine exonuclease) is the only described gene associated with apomixis in Boechera species. The centromere-specific histone H3 variant encoded by CENH3 gene is essential for cell division. Mutations in CENH3 disrupt chromosome segregation during mitosis and meiosis since the attachment of spindle microtubules to a mutated form of the CENH3 histone fails. This paper presents in silico characteristic of APOLLO and CENH3 genes, which may affect apomixis. Also, in this research we characterize the structure of CENH3, study expression levels of CENH3 and APOLLO in gynoecium/siliques of the natural diploid apomictic and sexual Boechera species at the stages of before and after fertilization. At the premeiotic stage, the expression level of CENH3 in the gynoecium of apomicts was two times lower than that of the sexual Boechera, it decreased in both species by the time of meiosis and increased after fertilization. By 1 DAP CENH3 expression started dropping in sexual B. stricta siliques and kept increasing in apomictic B. divaricarpa ones. That might indicate to a role of CENH3 in apomictic development in Boechera species. The expression levels of APOLLO also sharply decreased by the time of meiosis in gynoecium of both species; however, by 3 DAP, the level of APOLLO expression in siliques of apomicts was almost 1.5 times higher than that of the sexuals. While CENH3 was a single copy gene in all Boechera species, the APOLLO gene have several polymorphic alleles associated with sexual and apomictic reproduction in the Boechera genera. We also discuss polymorphism and phylogeny of the APOLLO and CENH3 genes.

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The functions and consequences of force at kinetochores

The Journal of Cell Biology ◽

10.1083/jcb.201211113 ◽

2013 ◽

Vol 200 (5) ◽

pp. 557-565 ◽

Cited By ~ 34

Author(s):

Florencia Rago ◽

Iain M. Cheeseman

Keyword(s):

Recent Work ◽

Chromosome Segregation ◽

Chromosome Movement ◽

Classical Studies ◽

Spindle Microtubules

Chromosome segregation requires the generation of force at the kinetochore—the multiprotein structure that facilitates attachment of chromosomes to spindle microtubules. This force is required both to move chromosomes and to signal the formation of proper bioriented attachments. To understand the role of force in these processes, it is critical to define how force is generated at kinetochores, the contributions of this force to chromosome movement, and how the kinetochore is structured and organized to withstand and respond to force. Classical studies and recent work provide a framework to dissect the mechanisms, functions, and consequences of force at kinetochores.

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CENP-C is a blueprint for constitutive centromere–associated network assembly within human kinetochores

The Journal of Cell Biology ◽

10.1083/jcb.201412028 ◽

2015 ◽

Vol 210 (1) ◽

pp. 11-22 ◽

Cited By ~ 87

Author(s):

Kerstin Klare ◽

John R. Weir ◽

Federica Basilico ◽

Tomasz Zimniak ◽

Lucia Massimiliano ◽

...

Keyword(s):

Cell Division ◽

Chromosome Segregation ◽

The Other ◽

Microtubule Binding ◽

Kinetochore Assembly ◽

Binding Interface ◽

Other Hand ◽

Spindle Microtubules ◽

Data Point

Kinetochores are multisubunit complexes that assemble on centromeres to bind spindle microtubules and promote faithful chromosome segregation during cell division. A 16-subunit complex named the constitutive centromere–associated network (CCAN) creates the centromere–kinetochore interface. CENP-C, a CCAN subunit, is crucial for kinetochore assembly because it links centromeres with the microtubule-binding interface of kinetochores. The role of CENP-C in CCAN organization, on the other hand, had been incompletely understood. In this paper, we combined biochemical reconstitution and cellular investigations to unveil how CENP-C promotes kinetochore targeting of other CCAN subunits. The so-called PEST domain in the N-terminal half of CENP-C interacted directly with the four-subunit CCAN subcomplex CENP-HIKM. We identified crucial determinants of this interaction whose mutation prevented kinetochore localization of CENP-HIKM and of CENP-TW, another CCAN subcomplex. When considered together with previous observations, our data point to CENP-C as a blueprint for kinetochore assembly.

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RANBP1 localizes a subset of mitotic regulatory factors on spindle microtubules and regulates chromosome segregation in human cells

Journal of Cell Science ◽

10.1242/jcs.009308 ◽

2007 ◽

Vol 120 (21) ◽

pp. 3748-3761 ◽

Cited By ~ 35

Author(s):

A. Tedeschi ◽

M. Ciciarello ◽

R. Mangiacasale ◽

E. Roscioli ◽

W. M. Rensen ◽

...

Keyword(s):

Chromosome Segregation ◽

Human Cells ◽

Regulatory Factors ◽

Spindle Microtubules

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Astrin-SKAP complex reconstitution reveals its kinetochore interaction with microtubule-bound Ndc80

eLife ◽

10.7554/elife.26866 ◽

2017 ◽

Vol 6 ◽

Cited By ~ 17

Author(s):

David M Kern ◽

Julie K Monda ◽

Kuan-Chung Su ◽

Elizabeth M Wilson-Kubalek ◽

Iain M Cheeseman

Keyword(s):

Chromosome Segregation ◽

Human Cells ◽

Cross Linking ◽

Microtubule Binding ◽

Binding Domains ◽

Anaphase Onset ◽

Ndc80 Complex ◽

Outstanding Question ◽

Dynamic Microtubule

Chromosome segregation requires robust interactions between the macromolecular kinetochore structure and dynamic microtubule polymers. A key outstanding question is how kinetochore-microtubule attachments are modulated to ensure that bi-oriented attachments are selectively stabilized and maintained. The Astrin-SKAP complex localizes preferentially to properly bi-oriented sister kinetochores, representing the final outer kinetochore component recruited prior to anaphase onset. Here, we reconstitute the 4-subunit Astrin-SKAP complex, including a novel MYCBP subunit. Our work demonstrates that the Astrin-SKAP complex contains separable kinetochore localization and microtubule binding domains. In addition, through cross-linking analysis in human cells and biochemical reconstitution, we show that the Astrin-SKAP complex binds synergistically to microtubules with the Ndc80 complex to form an integrated interface. We propose a model in which the Astrin-SKAP complex acts together with the Ndc80 complex to stabilize correctly formed kinetochore-microtubule interactions.

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Basic mechanism for biorientation of mitotic chromosomes is provided by the kinetochore geometry and indiscriminate turnover of kinetochore microtubules

Molecular Biology of the Cell ◽

10.1091/mbc.e15-06-0384 ◽

2015 ◽

Vol 26 (22) ◽

pp. 3985-3998 ◽

Cited By ~ 20

Author(s):

Anatoly V. Zaytsev ◽

Ekaterina L. Grishchuk

Keyword(s):

Error Correction ◽

Chromosome Segregation ◽

Basic Mechanism ◽

Spindle Pole ◽

Mitotic Division ◽

Human Cells ◽

Chromosome Loss ◽

Mitotic Chromosomes ◽

Optimal Regime ◽

Sister Kinetochore

Accuracy of chromosome segregation relies on the ill-understood ability of mitotic kinetochores to biorient, whereupon each sister kinetochore forms microtubule (MT) attachments to only one spindle pole. Because initial MT attachments result from chance encounters with the kinetochores, biorientation must rely on specific mechanisms to avoid and resolve improper attachments. Here we use mathematical modeling to critically analyze the error-correction potential of a simplified biorientation mechanism, which involves the back-to-back arrangement of sister kinetochores and the marked instability of kinetochore–MT attachments. We show that a typical mammalian kinetochore operates in a near-optimal regime, in which the back-to-back kinetochore geometry and the indiscriminate kinetochore–MT turnover provide strong error-correction activity. In human cells, this mechanism alone can potentially enable normal segregation of 45 out of 46 chromosomes during one mitotic division, corresponding to a mis-segregation rate in the range of 10−1–10−2 per chromosome. This theoretical upper limit for chromosome segregation accuracy predicted with the basic mechanism is close to the mis-segregation rate in some cancer cells; however, it cannot explain the relatively low chromosome loss in diploid human cells, consistent with their reliance on additional mechanisms.

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The ATAD2/ANCCA homolog Yta7 cooperates with Scm3HJURP to deposit Cse4CENP-A at the centromere in yeast

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1917814117 ◽

2020 ◽

Vol 117 (10) ◽

pp. 5386-5393 ◽

Cited By ~ 2

Author(s):

Sara Shahnejat-Bushehri ◽

Ann E. Ehrenhofer-Murray

Keyword(s):

Saccharomyces Cerevisiae ◽

Chromosome Segregation ◽

Histone H3 ◽

Nucleosome Assembly ◽

Direct Role ◽

Kinetochore Assembly ◽

Histone H3 Variant ◽

Assembly Factor

The AAA+ ATPase and bromodomain factor ATAD2/ANCCA is overexpressed in many types of cancer, but how it contributes to tumorigenesis is not understood. Here, we report that the Saccharomyces cerevisiae homolog Yta7ATAD2 is a deposition factor for the centromeric histone H3 variant Cse4CENP-A at the centromere in yeast. Yta7ATAD2 regulates the levels of centromeric Cse4CENP-A in that yta7∆ causes reduced Cse4CENP-A deposition, whereas YTA7 overexpression causes increased Cse4CENP-A deposition. Yta7ATAD2 coimmunoprecipitates with Cse4CENP-A and is associated with the centromere, arguing for a direct role of Yta7ATAD2 in Cse4CENP-A deposition. Furthermore, increasing centromeric Cse4CENP-A levels by YTA7 overexpression requires the activity of Scm3HJURP, the centromeric nucleosome assembly factor. Importantly, Yta7ATAD2 interacts in vivo with Scm3HJURP, indicating that Yta7ATAD2 is a cochaperone for Scm3HJURP. The absence of Yta7 causes defects in growth and chromosome segregation with mutations in components of the inner kinetochore (CTF19/CCAN, Mif2CENP-C, Cbf1). Since Yta7ATAD2 is an AAA+ ATPase and potential hexameric unfoldase, our results suggest that it may unfold the Cse4CENP-A histone and hand it over to Scm3HJURP for subsequent deposition in the centromeric nucleosome. Furthermore, our findings suggest that ATAD2 overexpression may enhance malignant transformation in humans by misregulating centromeric CENP-A levels, thus leading to defects in kinetochore assembly and chromosome segregation.

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Proper chromosome alignment depends on BRCA2 phosphorylation by PLK1

10.1101/265934 ◽

2018 ◽

Cited By ~ 4

Author(s):

Åsa Ehlén ◽

Charlotte Martin ◽

Simona Miron ◽

Manon Julien ◽

François-Xavier Theillet ◽

...

Keyword(s):

Chromosome Segregation ◽

Tumor Suppressor Protein ◽

Genome Integrity ◽

Phosphorylation Sites ◽

Docking Site ◽

Direct Role ◽

Chromosome Alignment ◽

Bona Fide ◽

Tetrameric Complex

SummaryThe BRCA2 tumor suppressor protein is involved in the maintenance of genome integrity through its role in homologous recombination. In mitosis, BRCA2 is phosphorylated by Polo-like kinase 1 (PLK1). Here we describe how this phosphorylation contributes to the control of mitosis. We identified two highly conserved phosphorylation sites at S193 and T207 of BRCA2. Phosphorylated-T207 is a bona fide docking site for PLK1 as illustrated by the crystal structure of the BRCA2 peptide bound to PLK1 Polo-box domain. We found that BRCA2 bound to PLK1 forms a complex with the phosphatase PP2A and phosphorylated-BUBR1. Reducing BRCA2 binding to PLK1, as observed in BRCA2 breast cancer variants S206C and T207A, alters the tetrameric complex resulting in misaligned chromosomes, faulty chromosome segregation and aneuploidy. We thus reveal a direct role of BRCA2 in the alignment of chromosomes, distinct from its DNA repair function, with important consequences on chromosome stability. These findings may explain in part the aneuploidy observed in BRCA2-mutated tumors.

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